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Relationship between serum antiheat shock protein 27 antibody levels and obesity
Article (Accepted Version)
http://sro.sussex.ac.uk
Kargar, Mehrdad, Tavassoli, Samira, Avan, Amir, Ebrahim, Mahmoud, Azarpazhooh, Mahmoud Reza, Asoodeh, Rasool, Nematy, Mohsen, Hassanian, Seyed Mahdi, Rahmani, Farzad, Mohammadzade, Elham, Esmaeili, Habibollah, Moohebat, Mohsen, Ferns, Gordon A, Ghayour-Mobarhan, Majid and Parizadeh, Seyed Mohammed Reza (2017) Relationship between serum anti-heat shock protein 27 antibody levels and obesity. Clinical Biochemistry, 50 (12). pp. 690-695. ISSN 0009-9120
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Accepted Manuscript
Relationship between serum anti-heat shock protein 27 antibodylevels and obesity
Mehrdad Kargari, Samira Tavassoli, Amir Avan, MahmoudEbrahimi, Mahmoud Reza Azarpazhooh, Rasool Asoodeh,Mohsen Nematy, Seyed Mahdi Hassanian, Farzad Rahmani,Elham Mohammadzade, Habibollah Esmaeili, MohsenMoohebati, Gordon A. Ferns, Majid Ghayour-Mobarhan, SeyedMohammad Reza Parizadeh
PII: S0009-9120(16)30324-1DOI: doi: 10.1016/j.clinbiochem.2017.02.015Reference: CLB 9481
To appear in: Clinical Biochemistry
Received date: 22 September 2016Revised date: 14 February 2017Accepted date: 14 February 2017
Please cite this article as: Mehrdad Kargari, Samira Tavassoli, Amir Avan, MahmoudEbrahimi, Mahmoud Reza Azarpazhooh, Rasool Asoodeh, Mohsen Nematy, Seyed MahdiHassanian, Farzad Rahmani, Elham Mohammadzade, Habibollah Esmaeili, MohsenMoohebati, Gordon A. Ferns, Majid Ghayour-Mobarhan, Seyed Mohammad RezaParizadeh , Relationship between serum anti-heat shock protein 27 antibody levels andobesity. The address for the corresponding author was captured as affiliation for allauthors. Please check if appropriate. Clb(2017), doi: 10.1016/j.clinbiochem.2017.02.015
This is a PDF file of an unedited manuscript that has been accepted for publication. Asa service to our customers we are providing this early version of the manuscript. Themanuscript will undergo copyediting, typesetting, and review of the resulting proof beforeit is published in its final form. Please note that during the production process errors maybe discovered which could affect the content, and all legal disclaimers that apply to thejournal pertain.
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Relationship between serum anti-heat shock protein 27 antibody levels and obesity
Mehrdad Kargari1,2,3,*
, Samira Tavassoli4,*
, Amir Avan3,5,*
, Mahmoud Ebrahimi6, Mahmoud
Reza Azarpazhooh6, Rasool Asoodeh
7, Mohsen Nematy
3, Seyed Mahdi Hassanian
1,3, Farzad
Rahmani1,3
, Elham Mohammadzade3, Habibollah Esmaeili
3,4, Mohsen Moohebati
6, Gordon
A. Ferns8, Majid Ghayour-Mobarhan
3,5, †, Seyed Mohammad Reza Parizadeh
3,†
Affiliations:
(1) Department of Clinical Biochemistry, School of Medicine. Mashhad University of
Medical Sciences, Mashhad, Iran; (2) Department of Basic Medical Sciences, Neyshabur
University of Medical Sciences, Neyshabur, Iran; (3) Metabolic syndrome Research Center,
School of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran; (4)
Department of Biochemistry, Payam Noor University of Mashhad, Mashhad, Iran; (5)
Department of Modern Sciences and Technologies, School of Medicine, Mashhad University
of Medical Sciences, Mashhad, Iran; (6) Cardiovascular Research Center, School of
Medicine, Mashhad University of Medical Sciences, Mashhad, Iran; (7) Department of
biostatistics and Epidemiology, School of Medicine, Mashhad University of Medical
Sciences, Mashhad, Iran; (8) Brighton & Sussex Medical School, Division of Medical
Education, Falmer, Brighton, Sussex BN1 9PH, UK.
†Corresponding Author:
Majid Ghayour-Mobarhan, MD, PhD; Metabolic syndrome Research Center, School of
Medicine, Mashhad University of Medical Sciences, Mashhad, Iran, 99199-91766,
Tel:+985118002288, Fax: +985118002287; Email: [email protected].
Seyed Mohammad Reza Parizadeh, PhD, Metabolic syndrome Research Center, School of
Medicine, Mashhad University of Medical Sciences, Mashhad, Iran, 99199-91766,
Tel:+985118002288, Fax: +985118002287; Email: [email protected].
Running title: Anti-HSP27 antibody levels in obese subjects
Grant: this study was supported by grant from Mashhad University of Medical Sciences
* Equally contributed as first authors
Conflict of interest: The authors have no conflict of interest to disclose
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Abstract
Background: Heat shock protein 27 (HSP27) is an intracellular molecular chaperone that is
expressed at high levels following the exposure of cells to environmental stressors such as
heat, toxins, and free radicals. High levels of HSP antigens and antibody titers have been
reported in several conditions including cardiovascular disease and cancers. We measured
serum anti-HSP27 antibody levels in 993 subjects and assessed the associations between
serum anti-HSP27 antibody levels and demographic characteristics including coronary risk
factors.
Methods: A total of 993 subjects were recruited as part of the Mashhad Stroke and Heart
Atherosclerotic Disorders (MASHAD) cohort study. Demographic, clinical, and biochemical
parameters and serum anti-HSP27 antibody titers were determined in all the subjects.
Results: Serum anti-HSP27 antibody levels increased with increasing age in men. No
significant differences in levels were detected between men and women. Serum anti-HSP27
antibody levels were significantly higher in obese subjects than in nonobese subjects
(P=0.046); however, no significant influence of smoking status was observed. Moreover,
serum anti-HSP27 antibody titers were positively associated with age, body mass index,
waist/hip ratio, the presence of diabetes mellitus, nonsmoking habit, serum triglycerides,
cholesterol, and high-sensitivity c-reactive protein.
Conclusion: We have found that serum anti-HSP27 antibody titers are related to several
cardiovascular risk factors, necessitating further studies on the value of this emerging marker
for risk stratification.
Keywords: Anti-heat Shock Protein 27, Cardiovascular Risk Factors, Determinants
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Introduction:
Heat shock protein 27 (HSP27) is an intracellular molecular chaperone that is expressed in
response to the exposure of cells to environmental stressors (1) and increases the ability of
cells to overcome environmental stress (2). HSPs modify several aspects of cell function,
including differentiation, apoptosis, and the repair and refolding of denatured proteins (3).
They are associated with cell exposure to a wide variety of environmental stresses including
high temperature, viral infection, oxidative stress, ischemia, heavy metals, ultraviolet
irradiation, and reactive oxygen species (ROS) (4).
HSPs are highly immunogenic, and it is believed that an autoimmune response to HSPs
contributes to rheumatoid arthritis, multiple sclerosis, inflammatory bowel disease, lupus
erythematosus (5, 6), and diabetes mellitus (1). There are several HSP families of proteins:
HSPH (former name HSP110), HSPC (HSP90), HSPA (HSP70), HSPD/E (HSP60/HSP10),
CCT (TRiC), DNAJ (HSP40), and HSPB (small HSP or sHSP) including HSP27 (7). HSP27
is a member of the sHSP family of proteins (8), and high serum titers of this protein have
been reported in individuals with cardiovascular disease (CVD) (9). HSP27 has been
suggested as a marker of inflammation (10), and immune response to HSPs may contribute to
the progression of atherosclerosis. Anti-HSP27 antibodies titers were increased during
atherogenesis (9, 11). Serum anti-HSP27 antibody titers have been suggested to predict
outcomes in cancer and may help to identify individuals at high risk of CVD (1).
There is growing evidence of a relationship between HSPs and inflammatory diseases,
including CVD. Most studies have focused on HSP/anti-HSP60, 65, and 70, although recent
data support the role of HSP/anti-HSP27 in CVD, which is the leading cause of death
worldwide (9, 12, 13).
We have previously demonstrated an association between hypertension and serum anti-
HSP27 antibody levels; however, the association with diabetes mellitus and smoking status
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was not statistically significant for a sample of 168 patients. Moreover, we previously
reported a relationship between anti-HSP27 antibody levels and risk of myocardial infarction
(13). The present study aimed to investigate serum anti-HSP27 antibody levels and determine
factors affecting it in a large population.
Materials and methods:
Population
A total of 933 subjects, including 456 men and 477 women, of age 35-64 years, were enrolled
as part of the Mashhad Stroke and Heart Atherosclerotic Disorders Study, Mashhad
University of Medical Sciences (14). Exclusion criteria included subjects with severe
hypertension, CVD or history of CVD, cerebrovascular disease, endocrine abnormalities,
infectious disease, diabetes mellitus, and pregnant or breastfeeding women (14, 15). The
protocol was approved by the Ethics Committee of Mashhad University of Medical Sciences,
and all the subjects provided written informed consent to participate in the study.
Anthropometric measurements
Anthropometric parameters including height, body weight, waist circumference, and hip
circumference were measured for all participants as described previously (15). Body mass
index (BMI) was calculated as weight (kg) divided by height squared (m2). In addition, we
collected information about the socioeconomic status of our subjects. The subjects were
divided into three subgroups: obese (BMI≥30 kg/m2), overweight (30>BMI≥25 kg/m
2), and
normal weight (BMI<25 kg/m2) (14, 15).
Biochemical analysis
Lipid profile, including total cholesterol, triglycerides, high-density lipoprotein cholesterol
(HDL-C), and low-density lipoprotein cholesterol (LDL-C), was evaluated in all the subjects.
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Fasting blood glucose (FBG) levels were measured as described before (14-15) using the BT-
3000 auto-analyzer machine (Biotechnica, Rome, Italy). High-sensitivity C-reactive protein
(hs-CRP) was measured by a PEG-enhanced immunoturbidimetry method using an Alycon
analyzer (ABBOTT, Chicago, IL, USA).
Serum anti-HSP27 antibody titer measurements
Anti-HSP27 antibody titers were determined in serum as previously described (16, 17).
Briefly, microtiter plates (Nunc Maxisorp, 3Nunc) were coated with 100 ng of recombinant
human HSP27 dissolved in 50 μL of carbonate buffer followed by incubation for 18 h at 4
°C. The wells were washed thrice with wash buffer (Phosphate-buffered saline (PBS)
containing 0.05% Tween-20). Nonspecific binding was reduced by blocking each well with
2% goat serum in PBS: 250 μL added to each well and incubated for 30 min at 37 °C and 30
min at room temperature. Serum was diluted to 1:100 with 2% goat serum in PBS. After
washing, 100 μL of peroxidase-conjugated goat anti-human IgG (Sigma-Aldrich, 4Inc., USA)
diluted to 1:500 with 2% goat serum in PBS was added to each well and incubated for 30 min
at room temperature. Subsequently, 100 μL of tetramethylbenzidine (TMB) substrate [200 μL
of 6 mg⁄mL TMB in Dimethyl Sulfoxide (DMSO) added to 10 mL of 50 mM acetate buffer,
pH 4.5, containing 6 μL of hydrogen peroxide (H2O2)] was added to each well, and the plate
was incubated for 15 min in the dark at room temperature. The reaction was terminated by
adding 50 μL of 2 M HCl to each well. Optical density was measured using a Labsystems
iEMS Reader Microtiter plate reader. After correction for nonspecific background absorbance
(subtracting the absorbance of uncoated wells from the antigen-coated wells for each
sample), the results were expressed in optical density units.
Statistical analysis
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Data were analyzed using SPSS software (version 11.5). The normality of distribution was
assessed using the Kolmogorov-Smirnov test. Descriptive statistics (mean±standard deviation
(SD) or median±interquartile range (IQR) were determined for normally or non-normally
distributed variables, respectively. For normally and non-normally distributed variables,
Student’s t-test and Mann-Whitney U test were used, respectively, to compare the clinical
characteristics and baseline demographics between the groups. Chi-square test and/or Fisher
exact test were used for comparing categorical variables. Pearson and Spearman correlation
tests were used to show significant correlation between two normal and two non-normal
quantitative variables, respectively. Non-normally distributed data were analyzed by
Kruskal–Wallis test (for more than two groups). Spearman correlation was used to investigate
the relationship between the anti-HSP antibody titers and atherosclerosis risk factors
including age, serum total cholesterol, hs-CRP, triglyceride, HDL-C, LDL-C,
cholesterol/HDL-C ratio, BMI, systolic and diastolic blood pressure, FBG, and waist and hip
circumferences, height, and waist/hip ratio. We determined the factors that were significantly
related to serum HSP27 antibody titers using stepwise multiple regression. A two-sided
P<0.05 was considered significant.
Results
Effect of gender, age, and smoking status on serum anti-HSP27 antibody levels
As shown in Table 1, age, FBG, waist/hip ratio, LDL-C, serum triglycerides, systolic blood
pressure, and presence of other CVD risk factors including diabetes mellitus and
hypertension (P>0.05) were not significantly different between the groups. However, women
had a significantly higher BMI, waist and hip circumferences, HDL-C, and hs-CRP (Table 1).
Serum anti-HSP27 antibody levels did not differ between men and women. Similar results
were also observed after dividing the participants in to age cohorts: 35-44, 45-54, and 55-65
years (Table 2).
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Effect of BMI on serum anti-HSP27 antibody level
Obese subjects had significantly higher BMI, waist/hip ratio, FBG, serum triglyceride, hs-
CRP, SBP/DBP, and LDL-C; HDL-C levels were lower in obese than in nonobese subjects
(P<0.05). Hypertension and dyslipidemia were more common in the obese subjects than in
the nonobese subjects (Table 3). Serum anti-HSP27 antibody titer was significantly higher in
the obese subjects than in the nonobese subjects (P=0.046, Table 5). After adjustment, the
differences between the groups remained significant (BMI: β=0.036, P=0.037).
Effect of socioeconomic status on serum anti-HSP27 antibody levels
Subjects were divided into six subgroups of educational attainment: primary school,
secondary school, high school, diploma, bachelor or postgraduate, and PhD or MD. Serum
anti-HSP27 antibody titers were not significantly different between the groups. Similar
results were observed for the effect of employment status on anti-HSP27 antibody levels
(Table 4-5).
Effect of atherosclerosis risk factors on anti-HSP27 antibody levels
Associations between anti-HSP27 antibody levels and atherosclerosis risk factors were
assessed by general linear model analysis as the dependent variable (anti-HSP27) is
quantitative and independent variables are both qualitative and quantitative. These results
showed that age, BMI, waist/hip ratio, triglycerides, cholesterol, hs-CRP, hyperlipidemia, and
hypertension were positively associated with serum anti-HSP27 antibody titers (Table 6).
Discussion
To the best of our knowledge, this is the first study evaluating the effect of demographic
characteristics and socioeconomic status with respect to serum anti-HSP27 antibody titers.
We found that obese subjects had significantly higher anti-HSP27 antibody titers than the
nonobese group, which is in agreement with our previous report (16). Therefore, our recent
data suggest that this biological marker can be used as a novel risk biomarker for CVDs.
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There is growing body of data suggesting the important role of this marker in CAD and
stroke (12-13, 16-18). In the present study, we also investigated its relationship with age. Our
data showed that serum anti-HSP27 antibody titer did not differ with gender, which is
consistent with the results of Zilaee et al, who observed no significant relationship between
gender and antibody titers to HSP27 and 60 (18). Moreover, Rea et al, in a study of 60
individuals aged between 20 and 96 years, showed that the anti-HSP70 antibody levels
increased significantly with age (19). Another study also reported that serum anti-HSP27
antibody levels increased with age, and it was associated with hypertension (20). We found a
significant increase in anti-HSP27 antibody levels with respect to age in the male subgroup,
which is in agreement with a previous report (20). Moreover, we explored the effect of
smoking status on serum anti-HSP27 antibody titers. However, no statistically significant
differences were observed between groups, which is consistent with our previous data (21).
There is accumulating evidence showing the association of inflammatory markers such as
CRP, TNF-α, and IL-6 with obesity (22). Our previous data revealed the association of body
mass and antibody titers to HSP60, 65, and 70 in 170 healthy Caucasians (23). In the current
study, we showed for the first time the important role of serum anti-HSP27 antibody titers as
a biomarker in obese subjects. Previous studies have shown that serum anti-HSP27 antibody
levels may reflect inflammatory status (10, 23). The role of anti-HSP antibody titers has been
shown during the process of atherosclerosis (9, 11), and the high levels of anti-HSP27
antibodies in obese subjects suggest that an immune response to HSPs contributes to the
progression of atherosclerosis in these subjects.
The effect of socioeconomic status on serum anti-HSP27 antibody titers was studied in our
population, although no statistically significant relationship was observed for these
parameters. Victora et al (2007) showed no association between serum anti-HSP60 antibody
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titers and maternal schooling, skin color, or any other socioeconomic or environmental status
(24).
A major strength of the present study is that it was performed in a large number of subjects,
and the effect of demographic, biochemical, and socioeconomic status of the population on
the anti-HSP27 antibody levels was studied. Moreover, this is the first study showing the
effect of obesity, a risk factor for CVD, on the level of serum anti-HSP27 antibody titers.
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Table 1: Comparison of clinical and biochemical characteristics between men and women
Total men women p-value
933 456 477 -
Age (year) 47.35±7.97 47.37±7.97 47.34±7.98 0.974
Height (m) 1.62±0.09 1.69±0.07 1.56±0.06 0.001
Weight (kg) 73.44±13.14 75.75±13.7 71.24±12.2 0.001
BMI (kg/m2) 27.92±4.75 26.39±4.32 29.39±4.69 0.001
FBG (mg/dL) 90.16±35.32 90.37±30.92 89.96±39.11 0.081
WC (cm) 95.58±12.25 93.49±11.07 97.56±12.98 0.001
HC (cm) 103.72±9.34 101.06±8.05 106.27±9.78 0.001
WHR 0.92±0.09 0.93±0.1 0.92±0.09 0.635
Serum LDL-C (mg/dL) 116.17±36.7 113.85±36.25 118.39±37.02 0.064
Serum HDL-C (mg/dL) 42.42±9.91 40.05±9.73 44.71±9.55 0.001
Serum TG (mg/dL) 116(84-168) 121 (87-172) 113 (81-162) 0.153
Serum hs-CRP (mg/dL) 1.61(1-3.46) 1.45 (0.89-3) 1.9 (1.11-3.7) 0.001
SBP (mmHg) 121.44±18.06 121.69±16.23 121.21±19.66 0.246
DBP (mmHg) 79.12±11 79.85±10.44 78.42±11.48 0.025
Diabetics (%) 7.45% (69.926) 7.51% (34.453) 7.4% (35.473) 0.771
Smoker (%) 24.44% (228.933) 29.61% (135.456) 19.5%(93.477) 0.001
Hypertensive (%) 22.07% (203.920) 22.27% (100.449) 21.87% (103.471) 0.883
Hyperlipidemic (%) 36.02% (335.930) 32.68% (149.456) 39.24% (186.474) 0.037
Anti-HSP 27 (OD) 0.23(0.1-0.43) 0.24 (0.11-0.43) 0.22 (0.1-0.43) 0.563
BMI, body mass index; FBG, fasting blood sugar; WC, waist circumferences; HC, hip circumferences; WHR,
waist/hip ratio; LDL-C, low-density lipoprotein cholesterol, HDL-C, high-density lipoprotein cholesterol; TG,
triglycerides; hs-CRP, high-sensitivity C-reactive protein; SBP, systolic blood pressure; DBP, diastolic blood
pressure.
Diabetes was defined as fasting blood glucose of ≥126 mg/L. Hypertension was defined as SBP of ≥140 mmHg
and DBP of ≥90 mmHg. Hyperlipidemia was defined as a total cholesterol ≥5.2 mmol/L (200 mg/dL),
triglycerides ≥1.5 mmol/L (150 mg/dL), and HDL-C <0.9 mmol/L (40 mg/dL) (for men) and <1.29 mmol/L (50
mg/dL) (for women).
Values are expressed as mean±SD, and interquartile range or median and Mann–Whitney U test were used for
normally and non-normally distributed data, respectively.
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Table 2: Comparison of clinical and biochemical characteristics between subgroups for age
Age (year) 35-44 45-54 55-65 p-value
number 393 349 191 -
Height (m) 1.63±0.09 1.62±0.09 1.6±0.1 0.004
Weight (kg) 74.08±14.45 72.82±12.11 73.26±12.1 0.783
BMI (kg/m2) 27.81±5.05 27.72±4.57 28.53±4.42 0.047
FBG (mg/dl) 85.73±27.18 91.85±35.74 96.17±46.61 0.001
WC (cm) 94.28±12.36 95.65±12.11 98.11±11.93 0.001
HC (cm) 104.2±9.74 103.34±9.12 103.47±8.95 0.53
WHR 0.91±0.11 0.93±0.08 0.95±0.07 0.001
Serum LDL-C (mg/dL) 109.81±36.14 118.83±35.94 124.46±37.15 0.001
Serum HDL-C (mg/dL) 41.84±9.68 43.4±10.54 41.85±9.08 0.118
Serum TG (mg/dL) 112 (77.25-157.75) 119 (84-166) 124 (99.75-186.25) 0.002
Serum hs-CRP (mg/dL) 1.54 (0.9-3.02) 1.62 (1.08-3.3) 2.01 (1.08-5.08) 0.006
SBP (mmHg) 115.11±14.53 123.23±17.21 131.19±20.91 0.001
DBP (mmHg) 76.52±10.76 80.25±10.67 82.39±10.93 0.001
Diabetics (%) 5.13% (20/390) 8.96% (31/346) 9.47% (18/190) 0.003
Smoker (%) 27.99% (110/393) 22.06% (77/349) 21.47% (41/191) 0.097
Hypertensive (%) 14.21% (55/387) 23.77% (82/345) 35.11% (66/188) 0.001
Hyperlipidemic (%) 28.75% (113/393) 38.9% (135/347) 45.79% (87/190) 0.001
Anti-HSP 27 (OD) 0.29±0.26 0.32±0.28 0.33±0.29 0.134
BMI, body mass index; FBG, fasting blood sugar; WC, waist circumferences; HC, hip circumferences; WHR,
waist/hip ratio; LDL-C, low-density lipoprotein cholesterol, HDL-C, high-density lipoprotein cholesterol; TG,
triglycerides; hs-CRP, high sensitivity c-reactive protein; SBP, systolic blood pressure; DBP, diastolic blood
pressure.
Diabetes was defined as fasting blood glucose of ≥126 mg/L. Hypertension was defined as a SBP ≥140 mmHg
and a DBP ≥90 mmHg. Hyperlipidemia was defined as a total cholesterol (TC) ≥5.2 mmol/l (200 mg/dl),
triglycerides (TG) ≥1.5 mmol/l (150 mg/dl), and HDL-C <0.9 mmol/l (40 mg/dl) (for men), and <1.29 mmol/l
(50 mg/dl)(for women).
Values are expressed as mean±SD or median(interquartile range) and One-Way ANOVA and Kruskal Wallis
test were used for normal and non-normal distributed data, respectively.
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Table 3: Comparison of clinical and biochemical characteristics between subgroups of BMI
p-value normal weight overweight obese
- 256 393 283 number
0.104 46.95±8.15 47.16±7.78 48±8.07 Age (year)
0.001 1.65±0.09 1.63±0.09 1.59±0.09 Height (m)
0.001 61.23±8.21 72.94±8.2 85.19±11.92 Weight (kg)
0.001 22.4±1.97 27.52±1.39 33.49±3.06 BMI (kg/m2)
0.006 87.72±40.5 89.46±30.3 93.14±36.48 FBG (mg/dl)
0.001 84.98±8.32 94.66±8.54 106.4±10.53 WC (cm)
0.001 95.41±5.01 102.51±5.9 112.9±8.15 HC (cm)
0.001 0.89±0.07 0.93±0.11 0.94±0.07 WHR
0.013 112.73±33. 115.21±38. 120.58±36.79 Serum LDL-C (mg/dL)
0.047 43.67±10.1 42.09±10.3 41.8±8.96 Serum HDL-C (mg/dL)
0.001 92.5 (66.75-125) 124 (89.25-178) 130(104.25-179.75) Serum TG (mg/dL)
0.001 1.22 (0.75-2.3) 1.51 (0.97-2.79) 2.81 (1.47-5.63) Serum hs-CRP (mg/dL)
0.001 117.02±16.0 121.37±17.0 125.52±19.64 SBP (mmHg)
0.001 76.19±10.7 79.18±10.5 81.68±11.15 DBP (mmHg)
0.033 5.1% 7.7% 8.9% (25/281) Diabetics (%)
0.714 25.39% 24.17% 24.03% (68.283) Smokers (%)
0.003 17.06% 21.13% (82/388) 27.86% (78/280) Hypertensive (%)
0.001 25.88% 39.03% 41.13% Hyperlipidemic (%)
0.046 0.23 (0.1-0.42) 0.21 (0.1-0.42) 0.27 (0.12-0.48) Anti-HSP27 (OD)
BMI, body mass index; FBG, fasting blood sugar; WC, waist circumferences; HC, hip circumferences; WHR,
waist/hip ratio; LDL-C, low-density lipoprotein cholesterol, HDL-C, high-density lipoprotein cholesterol; TG,
triglycerides; hs-CRP, high sensitivity c-reactive protein; SBP, systolic blood pressure; DBP, diastolic blood
pressure.
Diabetes was defined as fasting blood glucose of ≥126 mg/L. Hypertension was defined as a SBP ≥140 mmHg
and a DBP ≥90 mmHg. Hyperlipidaemia was defined as a total cholesterol (TC) ≥5.2 mmol/l (200 mg/dl),
triglycerides (TG) ≥1.5 mmol/l (150 mg/dl), and HDL-C <0.9 mmol/l (40 mg/dl) (for men), and <1.29 mmol/l
(50 mg/dl)(for women).
Values are expressed as mean±SD, and interquartile range or median and Mann–Whitney test were used for
normality and non-normality distributed data, respectively.
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Table 4: Comparison of clinical and biochemical characteristics between subgroups of educational attainment
p-
value PhD or MD
Bachelor or
Postgraduate Diploma High School
Secondary
School
Primary
School
- 6 69 27 362 358 110 Number
0.001 47.5±4.32 46.73±7.57 47.74±8.15 45.15±7.08 47.98±8 53.07±8.08 Age (year)
0.001 1.68±0.08 1.68±0.08 1.68±0.07 1.64±0.1 1.61±0.08 1.57±0.09 Height (m)
0.041 84.67±14.69 73.47±14.37 75.41±11.28 75±13.98 72.63±12.28 70.34±11.7 Weight (kg)
0.001 30.26±6.37 25.99±4.06 26.84±4.34 27.88±4.9 28.2±4.61 28.7±4.79 BMI (kg/m2)
0.153 113±50.63 88.55±38.16 90.38±29.19 88.37±32.29 90.5±30.22 95.36±54.78 FBG (mg/dL)
0.001 101±11.1 90.51±11.07 92.9±10.77 93.99±11.71 97.14±12.64 99.3±12.21 WC (cm)
0.045 107.08±11.67 100.4±7.19 102.09±6.97 103.62±9.9 104.46±9.24 104.06±9.11 HC (cm)
0.001 0.94±0.03 0.9±0.07 0.91±0.07 0.91±0.12 0.93±0.08 0.95±0.07 WHR
0.063 100.16±20.17 114.46±39.52 118.96±38.76 111.92±33.7 118.99±37.67 123.23±39.14 Serum LDL-C (mg/dL)
0.042 37.43±6.67 42.22±9.61 41.03±8.53 41.55±9.92 42.98±10.06 44.42±9.91 Serum HDL-C (mg/dL)
0.286 192.5 (114-
267.25)
114.5 (74.75-
170.75) 112 (63-176)
113 (83.75-
162) 118 (83-161)
128 (96.75-
178) Serum TG (mg/dL)
0.006 3.92 (1.42-
9.05)
1.18 (0.68-
2.32)
1.37 (0.77-
5.75)
1.51 (0.9-
3.21)
1.64 (1.06-
3.63)
1.9 (1.27-
4.29) Serum hs-CRP (mg/dL)
0.001 133.39±14.85 119.79±16.32 115.75±16.96 118.51±16.16 122.63±18.2 129.56±21.65 SBP (mmHg)
0.013 86.84±10.99 78.39±11.18 77.48±10.55 78.17±10.15 79.47±11.42 81.97±11.7 DBP (mmHg)
0.516 33.33% (2/6) 7.58% (5/66) 7.69% (2/26) 6.44%
(23/357)
8.15%
(29/356)
7.41%
(8/108) Diabetics (%)
0.245 33.33% (2/6) 18.18%
(12/66)
22.22%
(6/27)
20.89%
(75/359)
27.65%
(99/358) 30% (33/110) Smoker (%)
0.001 50% (3/6) 19.05%
(12/63)
25.93%
(7/27)
15.86%
(56/353)
24.23%
(86/355)
34.86%
(38/109) Hypertensive (%)
0.14 66.67% (4/6) 34.85%
(23/66)
37.04%
(10/27)
30.64%
(110/359)
38.94%
(139/357)
43.52%
(47/108) Hyperlipidemic (%)
0.295 0.23 (0.08-
0.59)
0.22 (0.08-
0.41)
0.2 (0.07-
0.45)
0.26 (0.13-
0.44)
0.21 (0.1-
0.43)
0.21 (0.09-
0.43) Anti-HSP 27 (OD)
BMI, body mass index; FBG, fasting blood sugar; WC, waist circumferences; HC, hip circumferences; WHR,
waist/hip ratio; LDL-C, low-density lipoprotein cholesterol, HDL-C, high-density lipoprotein cholesterol; TG,
triglycerides; hs-CRP, high sensitivity c-reactive protein; SBP, systolic blood pressure; DBP, diastolic blood
pressure.
Diabetes was defined as fasting blood glucose of ≥126 mg/L. Hypertension was defined as a SBP ≥140 mmHg
and a DBP ≥90 mmHg. Hyperlipidaemia was defined as a total cholesterol (TC) ≥5.2 mmol/l (200 mg/dl),
triglycerides (TG) ≥1.5 mmol/l (150 mg/dl), and HDL-C <0.9 mmol/l (40 mg/dl) (for men), and <1.29 mmol/l
(50 mg/dl)(for women).
Values are expressed as mean±SD, and interquartile range or median. Kruskal–Wallis and Chi-square tests were
used.
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Table 5: Comparison of clinical and biochemical characteristics between subgroups of employment status
p-value Retired Unemployed Employed
- 84 435 414 Number
0.001 54.65±6.46 47.53±8.02 45.69±7.32 Age (year)
0.001 1.66±0.09 1.56±0.07 1.68±0.08 Height (m)
0.001 74.57±12.32 70.57±11.68 76.24±14.12 Weight (kg)
0.001 27.06±3.96 28.9±4.67 27.07±4.8 BMI (kg/m2)
0.411 92.07±32.89 91.2±40.93 88.68±28.89 FBG (mg/dl)
0.012 94.09±11.13 96.85±12.86 94.53±11.69 WC (cm)
0.001 101.29±7.95 105.25±9.49 102.58±9.2 HC (cm)
0.39 0.93±0.07 0.92±0.9 0.92±0.11 WHR
0.507 114.55±38.39 117.43±35.5 115.18±37.61 Serum LDL-C (mg/dL)
0.001 41.85±10.31 43.98±9.46 40.91±10.06 Serum HDL-C (mg/dL)
0.273 122.5 (94-182) 113 (82-161.25) 120 (84-170) Serum TG (mg/dL)
0.02 1.59 (1.03-3.02) 1.84 (1.12-3.59) 1.44 (0.89-3.3) Serum hs-CRP (mg/dL)
0.011 126.49±16.62 121.41±20.11 120.44±15.76 SBP (mmHg)
0.026 82.33±9.71 78.61±11.69 79.01±10.4 DBP (mmHg)
0.96 8.33% (7/84) 7.89% (34/431) 6.81% (28/411) Diabetics (%)
0.014 16.67% (14/84) 21.84% (95/435) 28.74% (119/414) Smokers (%)
0.369 27.71% (23/83) 22.27% (96/431) 20.69% (84/406) Hypertensive (%)
0.436 34.52% (29/84) 38.19% (165/432) 34.06% (141/414) Hyperlipidemic (%)
0.183 0.28 (0.15-0.47) 0.23 (0.1-0.44) 0.23 (0.1-0.41) Anti-HSP27 (OD)
BMI, body mass index; FBG, fasting blood sugar; WC, waist circumferences; HC, hip circumferences; WHR,
waist/hip ratio; LDL-C, low-density lipoprotein cholesterol, HDL-C, high-density lipoprotein cholesterol; TG,
triglycerides; hs-CRP, high sensitivity c-reactive protein; SBP, systolic blood pressure; DBP, diastolic blood
pressure.
Diabetes was defined as fasting blood glucose of ≥126 mg/L. Hypertension was defined as a SBP ≥140 mmHg
and a DBP ≥90 mmHg. Hyperlipidemia was defined as a total cholesterol (TC) ≥5.2 mmol/l (200 mg/dl),
triglycerides (TG) ≥1.5 mmol/l (150 mg/dl), and HDL-C <0.9 mmol/l (40 mg/dl) (for men), and <1.29 mmol/l
(50 mg/dl)(for women).
Values are expressed as mean±SD, and interquartile range or median. Kruskal–Wallis and Chi-square tests were
used.
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Table 6: Association between anti-HSP27 antibody levels and risk factors of
atherosclerosis using general linear model analysis
Parameter Estimates
Dependent Variable: Anti_HSP27
Parameter β Std.
Error
t p-value
Intercept -1.901 0.974 1.951 0.051
Age 0.001 0.001 0.789 0.430
Height 1.216 0.587 2.072 0.039
Weight -0.012 0.006 1.948 0.052
BMI 0.037 0.017 2.210 0.027
WHR 0.100 0.103 0.972 0.331
Serum LDL-C -0.002 0.001 3.988 0.000
Serum HDL-C -0.002 0.001 1.648 0.100
Serum TG 0.000 0.000 3.620 0.000
Serum total cholesterol 0.002 0.001 2.842 0.005
Serum hs-CRP 0.001 0.001 0.687 0.492
Diabetes -0.040 0.036 1.123 0.262
Smokers -0.020 0.032 0.623 0.534
Hypertension 0.006 0.023 0.271 0.786
Hyperlipidemia 0.032 0.031 1.040 0.299
BMI, body mass index; WHR, waist/hip ratio; LDL-C, low-density lipoprotein cholesterol, HDL-C, high-
density lipoprotein cholesterol; TG, triglycerides; hs-CRP, high sensitivity c-reactive protein.
Diabetes was defined as fasting blood glucose of ≥126 mg/L. Hypertension was defined as a SBP ≥140 mmHg
and a DBP ≥90 mmHg. Hyperlipidemia was defined as a total cholesterol (TC) ≥5.2 mmol/l (200 mg/dl),
triglycerides (TG) ≥1.5 mmol/l (150 mg/dl), and HDL-C <0.9 mmol/l (40 mg/dl) (for men), and <1.29 mmol/l
(50 mg/dl)(for women).
General linear model analysis was performed as dependent variable (anti-HSP27) is quantitative and
independent variables are both qualitative and quantitative.
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Fig. 1
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Highlights
1. Serum anti-HSP27 antibody titers have an important role as biomarker in obese
subjects
2. Obese subjects had a significantly higher anti-HSP27 antibody levels than the
nonobese group
3. Serum anti-HSP27 antibody titers did not differ with gender
4. Serum anti-HSP27 antibody titers are related to several cardiovascular risk factors
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